Over 900 million people are at risk of acquiring lymphatic filariasis and it has been estimated that over 90 million are currently infected with Wuchereria bancrofti, Brugia malayi, or B. timori. The long-term objective of this study is to determine the molecular basis for protective immunity against filarial nematode parasites employing B. malayi as a model system. In these studies, we propose to focus our efforts on the proteins associated with the surface of the L3 and L4 stages of the parasite. Employing surface and metabolic labeling procedures and immunochemical analyses, we will characterize the composition and the time of synthesis of the major surface-associated components of the early infective forms of B. malayi. L3 surface-associated antigens will be characterized from larvae undergoing microfilaria-L2-L3 development in vivo in the mosquito vector. L4 surface-associated antigens will be studied in larvae undergoing L3-L4 development either in in vitro culture or in chambers implanted in a vertebrate host. The information on the composition and the interval of synthesis will be used to efficiently employ an innovative adaptation of the polymerase chain reaction (PCR) in a strategy to clone these surface constituents. In this adaptation of the PCR, the entire population of larval CDNAS are amplified in a procedure that results in the production of large amounts of larval PCRcDNAs all of which contain. restriction enzyme sites on the 5' and 3' ends so that the PCRcDNA products can be directionally cloned into a vector to form PCRcDNA libraries. Immunoscreening of PCRcDNA libraries will be carried out to identify and clone sequences encoding surface-associated molecules from B. malayi L3s and L4s. After characterizing the cloned DNAs and their protein products, the recombinant larval antigens will be expressed and used in a B. malayi-animal model system to test for their ability to induce protection against parasite challenge. In addition to providing information on the basic biology and biochemistry of the larval forms of this parasite, this study will provide recombinant reagents for the identification and study of the immune effector mechanisms that mediate protection against filarial infection.